Novel d-nor-progesterones and processes for their manufacture



United States Patent 3,352,920 NCVEL D-NGR-PROGETERGNES AND PROC- ESSES FUR THEHR MANUFACTURE Jerrold Meinwald, Ithaca, N.Y., assignor to Schering Corporation, Bioomrieid, N.J., a corporation of New Jersey N0 Drawing. Filed Feb. 25, 1965, Ser. No. 435,363 2 Claims. (Cl. 260-586) The rings are identified in the manner of conventional steroid nomenclature. Similarly, the carbon atoms of rings A, B and C are conventionally identified. In ring D, C17 is eliminated whereby C-16 is directly bonded to C-l3. The angular groups attached to positions and 13 retain conventional numbering and, as shown, R and R represent methyl or hydrogen.

In the conventional steroid nucleus, the six-membered C-ring is fused to thefive-membered D-ring in the trans configuration. The Dnor-steroids of the instant invention also possess the trans configuration between the C-ring and the now contracted D-ring.

Heretofore, steroids having a 4-membered cyclic D-ring were unknown. By my invention it is now possible to prepare a new class of 4-membered D-ring steroids (i.e., D-nor-Zl-desoxy pregnanes), which are characterized by being devoid of a 17-carbon atom and by having attached to the .16-carbon atom, moieties which are identical to those substituted at C-17 of a normal steroid possessing a S-membered D-ring, My D-nor-steroids include D-nor-Zldesoxy pregnanes, having at the 16-carbon atom a substituent possessing an oxygen function.

The D-nor-steroids have been found, in general, to possess therapeutic activities similar to the activities of the corresponding cyclopentyl-D-ring steroidal analog.

This invention thus provides a new class of steroids, i.e., D-nor-Zl-desoxy pregnanes, and, in particular, D-norprogesterones (D-nor-4-pregnene-3,ZO-diones) substituted at C-16 by a member selected from the group consisting of hydrogen, hydroxy, and lower alkanolyloxy, including the 19'-nor and l-dehydro analogs thereof.

Representative of the lower alkanoyl groups are radicals of lower alkanoic acids having preferably up to eight carbon atoms such as formic, acetic, propionic, butyric, caproic, valeric and enanthic acids.

This invention thus provides for the following specific D-nor-progesterones:

D-nor-progesterone (D-nor-4-pregnene-3,20-dione) D-nor- 1 -dehyd-roprogesterone; D-nor-ll9-nor-progesterone;

D-nor-l 6u-hydroxyprogesterone;

D-nor-l a-acetoxyprogesterone;

D-nor-16a-hydroxy-1-dehydroprogesterone and D-nor-l6a-acetoxy-1-dehydroprogesterone.

The above compounds are of value as progestational agents, for example, D-nor-l9-nor-progesterone exhibits progestational activity in rabbits via the intramuscular route when tested by the Clauberg assay at doses in the range of from about 0.8 rug/kg. to about 4 mg./kg., as well as oral progestational activity in rabbits at 20 mg./ kg. The above compounds are also valuable as intermediates in preparing useful steroids including other substituted D-nor-progesterones having progestational activity as discussed in greater detail hereinbelow.

It is to be understood that the above are given as exemplary, it being possible to prepare a D-nor-steroid corresponding to every known C-17 steroid by utilizing procedures known in the art for the normal cyclopentyl- D-ring steroids and substituting as starting compounds the corresponding D-nor-steroid.

The compounds of this invention, e.g., D-IIOI-llQ-IIOI- progesterone, may be administered orally or parenterally by incorporating a therapeutic dosage in conventional dosage forms such as tablets, capsules, elixirs, suspensions, solutions, or the like. They can be administered in admixture with pharmaceutical excipients which are chemically inert to D-nor-progesterone and derivatives thereof, such as, for example, cornstarch, lactose, sucrose, and gum arabic usually in admixture with an additive such as magnesium stearate, talc, and the like. The compositions may contain diluents and dispersing and surface active agents, and may be presented in a syrup, or in non-aqueous suspension, in aqueous suspensions, or in a syrup or in an oil.

The D-nor-progesterones are prepared from carboxy-D-nor-steroids such as D-nor-4-androstene-3-one- ISB-carboxylie acid and D-nor-S-androstene-3B-ol-1613- carboxylic acid by employing techniques analogous to methods known in the art for converting the carboxylic acid moiety in a C-17 carboxylic acid steroid (e.g., 5- androstene-3 3-ol-17,8-carboxylic acid) to a 17-acetyl, or other 17-substituents of known, normal, cyclopentyl-D- ring steroids.

The 16,8-carboxy-D-nor-steroids, necessary intermediates for the preparation of the novel D-nor-pregnanes of this invention, are prepared from l7-keto-androstanes and 17-ketoestranes unsubstituted in the 16-position via my novel process as described in US. Patent No. 3,113,142. In brief, my process is represented by the following reaction scheme A, wherein R represents the usual A, B and C-rings of a steroid:

0 ll W R N O H chloramine lower alkyl R n it ri t e by r "O0 aqueous solvent J 11-dione-16fi-carboxylic acid, D-nor-1,4,9(11)-androstadiene-3-one-l6fi-carboxylic acid, 9-flu0roD-nor-1,4-androstadiene11B-ol-3-one-1ofi-carboxylic acid, 6a-methyl- D nor-1,4-androstadiene-3,l1-dione-16li-carboxylic acid, a-fiuoro-D-nor-1,4-androstadiene 3,11 dione-l6fi-carboxylic acid and the like.

As mentioned heretofore, the D-nor-pregnanes of this invention are produced from 16-carboxy-D-nor-androstanes such as those listed above by utilizing transformations similar to those used in known conversions involving C-17 cyclopentanoid-Dring steroid structures. For example, D-norprogesterone is derived from 3,3-hydroxy-l6ficarboxy-D-nor-S-androstene which, after protection of the hydroxyl group at ,C3 by ester formation, is first converted to the corresponding lofi-acid chloride by treatment with a reagent such as thionyl chloride, phosphorus trichloride, phosphorus pentachloride, and preferably oxalyl chloride. The thus formed acid chloride, e.g., 3p?- acyloxy-l6 3-carbonyl chloride-D-nor--androstene, is in turn treated with an alkylating agent such as dimethyl cadmium, giving rise, after hydrolysis of the ester at C3, to the intermediary D-nor-5-pregnene-3,8-ol-Z0 one which, upon oxidation with chromic acid sulfuric acid reagent, yields D-nor-progesterone. Similarly,when S'fl-hydroxy- 16,8-carboxy-D-nor-l9-nor-5-androstene is subjected to the aforedescribed sequence of reactions, there is obtained D.-nor-19-nor-progesterone.

Alternatively, D-nor-4-androstene-3-one-16;8-carboxylic acid (prepared by the ultraviolet irradiation of 16-diazo- 4-androstene-3,17-dione, in turn derived from 16-diazo- 5-.androstene-3B-ol-17-one by the action of Flavobacterium dehydrogenans) is converted to the corresponding acid chloride by the action of oxalyl chloride, followed by reaction with diazomethane, and subsequent reaction of the thereby formed 2l-diazo-D-nor-4-pregnene-3,20- dione with hydrogen bromide followed by reaction of the resultant 2l-bromo compound with sodium iodide followed by acetic acid to give D-nor-pregesterone.

The 16a-hydroxyl groups are introduced into the D- nonpregnane molecule by techniques such as those whereby 21-diazo-D-nor-4-pregnene-3,20'dione (prepared 'as described in Example 1) is converted to D-nor-4-pregnene-16a,21-diol-3,20-dione. Conversion of this latter compound to 16a-hydroxy-D-nor-progesterone is easily effected by esterifying at C-Zl with methanesulfonyl chloride followed by reduction of the 2l-ester thereby formed by means of sodium iodide in acetone followed by treatment with acetic acid. Esterification of the 16cchydroxy group in 16a-hydroxy-D er-progesteroneis conveniently effected by reaction with an acid such as acetic or caproic in the presence of trifiuoroacetic anhydride whereby there is formed the 16-acetate and lo-caproate, respectively, of 16a-hydroxy-D-nor-progesterone.

In addition to being useful for protecting hydroxyl groups in processes such as are described herein, the esters of my D-nor-steroids are useful in that the therapeutic activities thereof are generally of longer duration than the corresponding hydroXy-D-noresteroid.

Introduction of a n -double bond into my novel D-norpregnanes possessing a 3-keto-A -system'may be effected by known chemical methods such as by means of selenium dioxide or by the use of dichlorodicyanobenzoquinone or alternatively, by microbiological methods utilizing microorganisms such as Corynebacterium simplex (A.T.C.C. 6946). For example, D-nor-progesterone, 16a-hydroxy- D-nor-progesterone (prepared as described hereinabove) are subjected to the action of Corynebacterium simplex according to procedures described in U.S. Patent No. 2,837,464 for conversion to D-nor-l-dehydro-progesterone (D nor 1,4 pregnadiene-3,20-dione) and l6a-hydroxy-1-dehydro-D-nor-progesterone (D nor-1,4-pregnadiene-16a-ol-3,20-dione) respectively.

The. A -4,5-dihydro-D-nor-pregnene analogs of D-nor compounds may be obtained from the corresponding 3-keto.-.D-nor 4-.pregnanes. For example, D-.nor-progesterone subjected to the action of hydrogen in the presence of a catalyst such as palladium in a solvent such as ethanol, ethyl acetate or tetrahydrofuran yields D-nor- 5u-pregnane-3,20-dione. Reaction of the aforementioned D-nOr-Su-pregnane with one equivalent of a halogen such as bromine yields the respective 2-bromo-analog which is dehydrohalogenated with a reagent such as collidine or dimethylformamide in the presence of calcium carbonate yielding D-nor-Sog-1-pregnene-3,20-dione.

It is to be understood that in the specification and in the examples, the conversions shown are by way of illustration only, it being obvious to one skilled in the art that analogous transformations may be carried out when other intermediates are used.

In general, the following additional groups will not interfere with the processes of this invention:

Hydroxy or acyloxy groups at one or more of positions 6, 11 and 15;

Keto groups at position 11;

Lower alkyl groups containing up to 4 carbon atoms and preferably methyl at one or more of positions such as at C-1, 2, 6 and 15;

Halogeno group, and particularly fluoro and chloro at one or more of positions C-6, 9, l1, and 15;

And unsaturated linkages.

When preparing D-nor steroids of this invention which are substituted at one or more of carbon atoms 2, 4, 6, 9, 11 and 15, it is preferable, as described hereinabove, to have all the substituents mentioned above in the 1613- carboxy-D-nor-steroid starting intermediate prior to building up the (3-16 pregnane side chainaHowever, substituents may be introduced in the molecule at any point during the preparation of the D-nor-pregnanes, by utilizing procedures known in the art.

For example, 9a,l1,6 dihalogeno-D-nor-progesterones are prepared by utilizing techniques on my D-n0r-9(11)- dehydroprogesterones similar to those described for the 9(11)-dehydro derivatives of normal, cyclopentyl D-ring steroids in US. Patents Nos. 2,894,963 and 3,009,928- 3,009,953. When a 911,11fi-dihalogeno-D-nor-progesterone is desired, the requisite 9(11)-dehydro intermediate is prepared by introducing an llfi-hydroxyl group into a D- nor-progesterone, such as 16a-acetoxy-D-nor-progesterone, by the action of Curvularia lunata followed by treatment of the 11,8,16a-dihydroxy-D-nor-progesterone 16-acetate thereby formed with methanesulfonyl chloride in dimethylformamide to give 16a-acetoXy-D-nor-9(11)- dehydroprogesterone. Reaction with chlorine in acetic acid according to known techniques yields 9a,1l;8-di-.

chloro-16rx-acetoxyD nor-progesterone which, in addition to possessing progestational activity isvaluable as a topical anti-inflammatory agent.

Substituents such as oc-hYdIOXY, a and B-lower alkyl, and a and fi-halogeno may be introduced into the D-nor steroid nucleus at C15 utilizing procedures similar to known methods for the introduction of these moieties at C-16 of a C-17 cyclopentanoid-D-ring steroid. For example, to introduce a ISa-rnethyl group into D-nor-pregneneolone (D-nor5-pregnene-3,8-ol-20-one) (prepared as described in Example 1D) the requisite IS-dehydro intermediateis first prepared by brominating D-nor-pregneneclone in chloroform with bromine, followed by treatment of the 5,6,16-tribromo derivative thereby formed with sodium iodide in acetone to give 16-bromo-D-nor-5- pregnene-35-ol-20-one which is heated with collidine to give the desired D-nor-5,1S-pregnadiene-3fi-ol-20-one. In similar manner, when D-nor-pregnane-3 8-o1-20-one is brominated then dehydrobrorninated as described above, there is obtained D-nor-l5-pregnene-3B-ol-20-one. Addition of a standard Grignard reagent, such as methyl magnesium iodide to a IS-dehydro-D-nor-steroid utilizing known techniques will result in the production of the corresponding 15oc-1'nethyl derivative, e.g., ISoc-IHCthYl-D- nor-5-pregnene-3fl-ol-ZO-one.

When other 15u-lower alkyl derivatives are desired,

other corresponding lower alkyl Grignard reagents are employed, e.g., ethylmagnesium bromide and isopropyl magnesium iodide or t-butyl magnesium iodide, whereby is obtained the corresponding la-ethyl-, l5a-isopropyl-, and ISu-t-butyl derivatives of D-nor-S-pregnene-3,B-ol-- one.

One method of introducing a 15,8-lower alkyl group into a D-nor-steroid such as D-nor-Sm-l5-pregnene-3l3-ol- 20-0ne, involves reaction of the lS-dehydro-bond with diazomethane whereby is obtained 15,16-pyrazoline-D- nor-5ot-pregnane-3/3-ol-20-one which, after pyrolysis at or above the melting point of the pyrazoline intermediate yields the corresponding IS-methyl l5 dehydro-llnorsteroid, e.g., l5-methyl-D-nor-5a-l5-pregnene-3fi-ol-20- one. Although it is preferred to pyrolyze by heating the pyrazoline above its melting point, such pyrolysis may be eifected by heating the substance in a high boiling, inert solvent, such as p-cymene, or tetraline or the like. The unsaturation at C-lS in the D-nor-ring is conveniently removed by reductive hydrogenation preferably in the presence of a catalyst, such as palladium, whereupon there is obtained 155-methyl-D-nor-5a-pregnane-Sfl-ol- ZO-one 3-acetate. In a similar manner, D-nor-5,l5-pregnadiene-3,8-ol-20-one upon reaction with diazomethane followed by pyrolysis and subsequent partial hydrogenation, yields l5,B-methyl-D-nor-S-pregnene-3fl-ol-20-one. Where the ISB-ethyl, 15,8-propyl, or 15,8-butyl derivative is desired, the corresponding diazoethane, diazopropane, diazobutane, or the like is used in the aforedescribed procedure.

The 15-alkyl-l5-dehydro intermediates prepared as described above are also valuable intermediates in the preparation of novel l5-methylene-D-nor-steroids. Thus, utilizing known techniques, 15-methyl-D-nor-5,l5-pregnadiene- 3/3-ol-20-one upon treatment with alkaline hydrogen peroxide yields the corresponding l5a,l6ot-epoxide, i.e., 15a, 16a oxide 15,8 methyl D nor 5 pregnene 3,8- ol-ZO-one. Treatment of the 15a,l6oc-OXidO-D-11OI-St6fl0d with a hydrogen halide, e.g., hydrogen bromide produces the corresponding l5-methylene-l6u-l1ydroxy derivative, e.g., l5-methylene-D-nor-5-pregnene-3{3, l 6[3-diol-20-one. The action of the microorganism Flavobacterium dehydrogenans utilizing conventional techniques will convert 15 methylene-D-nor-S-pregnene-3{3,l6a-diol-20-one to 15- methylene-l6a-hydroxy-D-nor-progesterone (lS-methylene-D-nor-4-pregnene-l6a-ol-3,20-dione) which may be acetylated with acetic anhydride and p-toluene sulfonic acid to give 15-methylene-16a-acetoxy-D-nor-progesterone, which is a potent progestational agent via the oral route.

From the 1S-methylene-l6a-hydroxy-D-nor-progesterones produced as described hereinabove, there may be obtained valuable 2l-oxygenated IS-methylene pregnanes possessing cortical activities. Thus, for example, 15- methylene-16a-hydroxy-D-nor-progesterone is transferred by oxidation with Rhizopus nigricans into the corresponding lla-hydroxy derivative, which, in turn, is transformed by the action of iodine/calcium oxide into the 2l-iodo derivative, transformed by acetolysis into the 21- acetoxy derivative, 15-methylene-D-nor-4-pregnene-lla, 16a,2l-triol-3,20-dione 2l-acetate. The conversion of the lla-hydroxy system to the corresponding 9u-fiuoro-ll5- hydroxy system follows methods similar to those well known in the art as described heretofore whereby is obtained 9a-fluoro-15-methylene-D-nor-hydrocortisone 2l-acetate, which on incubation with Corynebacterium simplex gives 9ot-fluoro-15-methylene-D-nor-prednisolone, a powerful corticoid, valuable as an anti-inflammatory. Alternatively, treatment of 1S-methylene-16a-hydroxy-D- nor-progesterone with the microorganism Curvularia lunata gives the corresponding llfi hydroxy derivative, 15 methylene D nor 4 pregnene 11,8,16a diol- 3,20-dione which after treatment with iodine/calcium oxide to the 21-iodo derivative followed by acetolysis yields lS-methylene-D-nor-hydrocortisone Zl-acetate; the

3 latter is converted to the l5-rnethylene-D-nor-prednisolone by Corynebacterium simplex.

The 15-methylene-D-nor-steroids prepared as described hereinabove are convertible by reduction into the corresponding 150aand lSB-methyl analogs. Thus, for example, IS-methylene-l6a-hydroxy-D-nor-progesterone upon reduction with one equivalent or" hydrogen with palladium on charcoal as catalyst yields a mixture of the lilac-methyl and lSfit-methyl isomers of l5-II16lllY1-16ochydroxy-D-nor-progesterone which are each separated utilizing chromatographic techniques whereby there is obtained 1Sa-methyI-l6a-hydroxy-D-nor-progesterone and 1SB-methyl-l6a-hydroxy-D-nor-progesterone.

The lS-methyl-D-nor-corticoids of my invention may alternatively 'be prepared from the IS-methyl-D-norpregnenolones, i.e., l5a-methyl-D-nor-5-pregnene-3fl-ol- ZO-one and l5B-methyl-D-nor-5-pregnen-3fl-ol-20-one by conversion to the 15-methyl-D-nor-progesterones via Oppenauer oxidation, hydroxylation with Glomerella cingu- [am or Rr'tizopus rtigricans according to known procedures to give 15a-rr1ethyl-lla-hydroxy-D-nor-progesterone and 15 ,B-rnethyll la-hydroxy-D-r1or-progesterone, respectively. The latter compounds are oxidized to the ll-ketones with chromic acid and then carried through the sequence of reactions for the elaboration of the corticoid structures according to procedures outlined in I. Am. Chem. Soc. 77: 4436 (1955) to give l5a-methyLD-nor-prednisolone and lSfi-methyl-D-nor-prednisolone respectively.

Still alternatively, l5ot-methyl-9a-fluoro-D-nor-prednisolone may be prepared from ISa-methyl-D-nQr-S-pregnene-3 3-ol-20one according to procedures outlined in I. Am. Chem. Soc. 80: 4431 (1958) for the preparation of 16a-methyl-9a-fiuoro-prednisolone.

The introduction of a lSa-hydroxyl group in my novel l5 unsubstituted-D-nor-steroids is conveniently effected utilizing microorganisms such as Hormodendrum olivacewm. (A.T.C.C. 13, 596), Collectotrichum antirrhea, Penicillium notwla, Caloneclria decora (by methods disclosed in German Patent 1,067,020) Gibberella baccata and Gibberella saubineth. A preferred method is to introduce the ISa-hydroXy group via Hormoaendrum olivaceum (A.T.C.C. 13, 596) according to procedures similar to those described for a Cl7 cyclopentyl-D-ring steroid by S. Bernstein et al. J. Am. Chem. Soc. 82: 3685 (1960).

The process of this invention may be used in the preparation of the following specific D-nor-progesterones and derivatives thereof as disclosed herei D-nor-progesterone D-norl 9-nor-progesterone; 6cr-chloro-D-nor-progesterone; 16e-liydroxy-D-nor-progesterone; l6wacetoxy-D-nor-progesterone;

l6a-capro oxy-D-nor-progesterone l 6a-hydroxy-D-norl -dehydroprogesterone; 16ot-acetoxy-D-nor4-dehydro-progesterone;

The following are examples which illustrates my invention. It is to be undestood that the invention is not to be limited to the exact details of operation or exact compounds shown and described, as obvious modifications and equivalents will be apparent to one skilled in the art; the invention is therefore to be limited only by the scope of the appended claims.

EXAMPLE 1 D-nor-progesierone- (D-n0r-4-pregnene-3,20-a'ione) A. D-NOR-S-A-NDROSTENE-3B-OL-l6B-CARBOXYLIC ACID CHLORIDE 3-ACETATE To 3 g. of D-nor-5-androstene-3fl-ol-l6fi-carboxylic acid 3-acetate in 25 ml. of anhydrous benzene acid 2.5 ml. of oxalyl chloride and heat the mixture under reflux for one hour. Distill the benzene in vacuo under anhydrous conditions. The resultant residue of substantially D-nor-S- androstene-3fi-ol-l6,6-carboxylic acid chloride 3-acetate is 7 used without further purification in the procedure immediately following.

B. 21DIAZO-DsNOR-S-PREGNENE3/3-OL20-ONE S-A'CTATE To a solution of 1 g. of D-nor--androstene-3 3-ol-16B- carboxylic acid chloride 3-acetate in 25 ml. of benzene add a solution of diazomethane in ether until a persistent yellow color is obtained. Allow the mixture to stand at room temperature for 30 minutes, then concentrate under reduced pressure to a residue of 21-ddiazo-D-nor-5-pregnene-3fl-ol-20-one 3-acetate. Purify by crystallization from ether-hexane.

C. D-NOR5-PREGNENE3B-OL 20-ONE S-ACETATE To a solution of 500 mg. of 21-diazo-D-nor-5-pregnene-3B-ol-20-one 3-acetate in 50 ml. of ether add a solution of about 500 mg. of hydrogen bromide in 15 ml. of ether. Stir the mixture at room temperature for 30 minutes then wash the solution with cold water, cold sodium bicarbonate solution and again With Water, then dry over magnesium sulfate and concentrate under reduced pressure. To the resultant residue containing 21- bromo-D-nor-5-pregnene-3B-ol-ZO-one 3-acetate add 15 ml. of acetone and a solution of 1.5 g. of sodium iodide in 15 ml. of' acetone. Heat the mixture at reflux temperature for one hour then add 1 ml. of acetic acid and heat the mixture under reflux for an additional 30 minutes. Decolorize the resulting brown solution by the addition of aqueous sodium bisulfite then pour the mixture into ice water. Filter the resulting precipitate, wash with water and dry to give D-nor-5-pregnene-3B-o1-20-one l t-acetate. Purify by crystallization from hexane.

Alternatively, the compound of this example is prepared as follows:

Prepare two equivalents of dimethyl cadmium in benzene according to the procedure in F. A. Shirley, Organic Reactions, vol. VIH, John Wiley and Sons, Inc., New York, NY. (1954). Add this benzene solution of dimethyl cadmium slowly and dropwise to a well stirred solution of D-nor-5-androstene-3fl-ol-16/3-carboxylic acid chloride 3-acetate prepared as described (in Example 1A), in 50 ml. of dry benzene. Stir the resultant mixture for 1 hour and then reflux for 15 minutes.

Acidify the reaction mixture then extract with ether. Combine the ethereal extracts, Wash with 3 N hydrochloricacid, water, then with sodium bicarbonate solution, and dry over magnesium sulfate. Evaporate the ether, and chromatograph the resultant residue over Woelm neutral alumina, activity grade III, eluting with 7:3 hexane-benzene. Combine the eluates and evaporate to a residue of D-nor-S-pregnene-3B-ol-20-one 3-acetate. Purify by recrystallization from hexane.

D. DNOR-5-PREG'NENE3B-OL-20-ONE Dissolve 300 mg. of D-nor-5-pregnene-3;8-ol- 20- one 3-acetate in a mixture of 10 ml. of acetone and 20 ml. of 5% hydrochloric acid in 90% aqueous methanol. Allow the solution to stand at room temperature for 24 hours then remove about half the solvent under reduced pressure. Pour the residual solution into ice water, filter the resultant precipitate of D-nor-5-pregnene-3fl-ol. Purify by crystallization from acetone-hexane.

E. D-NOR-PROGESTERONE To a solution of 250 mg. of D-nor-5-pregnene-3,8-ol- 20-one in 10 ml. of acetone chilled to 5 C., add dropwise chromic acid-sulfuric acid reagent (266 mgm. CrO ml.) until a permanent orange color is obtained. Keep the solution at room temperature for 5 minutes then add a little methanol to destroy any excess reagent. Pour the solution into ice Water and extract with ether. Combine the ether extracts, wash, dry over magnesium sulfate, and evaporate in vacuo to a residue of D-nor-progesterone. Purify as follows:

Dissolve the residue in acetone to which has been added about 2 ml. of 3 N hydrochloric acid. Allow the solution to stand at room temperature for 1 hour then pour into water. Filter the resultant precipitate, wash with water, dry, and crystallize from acetone-hexane.

EXAMPLE 2 Alternate procedure for the preparation of D-norprogesterone A. 1GDIAZO 4-ANDROSTENE-3,l7-DIONE Yeast extract (Difco) gm 10 Potassium phosphate monobasic gm 4.48 Sodium phosphate dibasic gm 4.68

Tap water to 1 liter.

This culture medium has previously been autoclaved, at 15 lb. pressure, for twenty minutes to obtain aseptic conditions, and cooledJThe variant is grown in the medium under constant illumination, using the visible range of the spectrum. The incubation temperature is maintained at about 33 C. and is conducted under aerobic conditions. Aeration is accomplished by agitation and/ or blowing air through the culture medium.

After the organism has grown for 12 to 24 hours (or longer, if desired), ml. of the growing culture are. introduced into each of ten flasks, and to each flask are added 200mg. of 16-diazo-5-androstene-3 3-ol-17-one dissolved in a minimum volume of ethanol. The reaction mixtures are then shaken at 30 C. for 12 to 72 hours. The reaction is stopped when paper chromatography indicates that there is no more starting material.

The contents of the flasks are combined and extracted with methylene chloride. The extracts are concentrated and the residue is crystallized from acetone-hexane yielding 16-diazo-4-androstene-3,17-dione.

B. DNOR-4-ANDROSTENE-3-ONE-1SBCARBOXYLIC ACID Irradiate a solution of 5 g. of l6-diazo-4-androstene- 3,17-dione in 5 ml. of aqueous dioxane for 4 hours with a 200 watt Mercury lamp provided with a Corax sleeve. Remove the solvent under reduced pressure and triturate the resultant residue with water and dry giving D-nor-4-' androstene-3-one-l6fl-carboxylic acid. Purify by crystallization from acetone.

C. D-NOR-4-A NDROSTENE-3-ONE-l6/ELCARB OXYLI C ACID CHLORIDE D. 21-DIAZO D-NOR-4-PREGNENE 3,ZO-DIONE To a solution of the D-nor-4-androstene-3-one-l6B- carboxylic acid chloride, prepared in Example 2C, in 20 ml. of benzene, add an ethereal solution of diazomethane until a persistent yellow color is obtained. Keep the mixture at room temperature for 30 minutes then concentrate in vacuo to a residue substantially of 21-diazo-D-nor-4- pregnene-3,20-dione. Purify by crystallization from acetone-hexane.

E. D-NOR4PREGNE-NE-3,20-DIONE EXAMPLE 3 D-nor-I ,4-pregnadiene-3,20-dione D-nor-4-pregnene-3,ZO-dione is subjected to the action of a culture of Corynebacterium simplex (A.T.C.C. No. 6946) in the following manner:

A solution of 1 g. of yeast extract (Difco) in one liter of tap water, the pH of which is adjusted to 6.9, is distributed among ten 300 ml. Erlenmeyer flasks and to each flask is added a loopful, 2 ml., of Corynebacterium simplex. The resulting suspensions are incubated at 30 C. on a shaking machine for 18 hours. One-half gram of D-nor-4-pregnene-3,20-dione is dissolved in 25 ml. of acetone and the resulting solution is distributed equally among the ten flasks containing the 18-hour growth of C. simplex. The culture containing the D-nor-4-pregnene- 3,20 dione is then incubated at 30 C. for 24 hours. At the end of 24 hours, the contents of the flasks are cornbined and extracted with a total of 3 liters of chloroform. The crude chloroform extract from the transformation is then conc;ntrated to a residue which is crystallized from methylene chloride-hexane, affording D-nor-1,4- pregnadiene-SJQdiOne.

EXAMPLE 4 6a-chl0ro-Dn0r-4-pregnene-3,20-di0ne A. 5,6DICHLOROD-NORPREGNANE-35-OL-20-O NE To a solution of 1 g. of D-nor-5-pregnene-3/3-ol-20- one (the compound of Example 1D) in a mixture of 25 ml. of carbon tetrachloride and 5 ml. of methylene chloride containing 0.75 ml; of pyridine add at -20 C. a solution of 240 mg. of chlorine in 2.7 ml. of carbon tetrachloride. Stir the mixture at 20 C. for 30 minutes then allow to warm to room temperature over a period of 30 minutes. Dilute the reaction mixture with methylene chloride and wash the organic solution with water, sodium thiosulfate solution, and again with water. Dry the methylene chloride solution with magnesium sulfate then concentrate to a residue of substantially 5,6-dich1oro-D-norpregnane-B/i-ol-ZO-one. Purify by crystallization from methylene chloride-pentane.

B. 5,G-DICHLORO-D-NOR-PREGNANE-3,20-DIONE Oxidize 5,6-dichloro-D-nor-pregnane-3fi-ol 2O one with chromic acid-sulfuric acid reagent according to the procedure of Example 1E. Isolate the resulting product in the described manner yielding 5,6-dichloro-D-nor-pregnane-3,20dione.

C. Ga CHLORO-D-NOR-ei-PREGNENE-3,2O-DIONE Pass a stream of hydrogen chloride through a solution of 150 mg. of 5,6-dichloro-D-nor-pregnane-3,ZO-dione in 20 ml. of acetone chilled to C. for 1 hour. Keep the reaction mixture at 0 C. for 2 hours longer than concentrate in vacuo to a residue of about ml. Pour the residue into water and filter the resulting precipitate, wash With water, and dry yielding substantially Gzx-ChlOIO-D- nor-4-pregnene-3,20-dione. Purify by crystallization from acetone-hexane.

10 EXAMPLE 5 D-n0r-4-pregnene-1 61:,2] -di0l-3,20-di0ne A. 21,21-DIBR OMO-D-N OR-4-PREGNENE-3,20-DI ONE To a stirred solution of 5 g. if 21-diazo-D-nor-4-pregnene-3,20-dione (the compound of Example 1B) in ml. of chloroform add dr-opwise a solution of 2.5 g. of bromine in 15 ml. of chloroform. Continue stirring until the bubbling has stopped and the bromine color has been discharged. Remove the solvent at room temperature under reduced pressure to a residue of substantially 21,21- dibromo-D-nor-4-pregnene-3,ZO-dione which is used without further purification in the procedure immediately following.

B. METHYL D-NOR-4,16 (20) -PREGNADIE NE3-ONE- 21-OATE To a solution of 2 g. of 21,2l-dibromo-D-nor-4-pregnene-3,20-dione in 40 ml. of methanol add 15 ml. of an 8% solution of sodium methoxide in methanol. Stir the mixture under nitrogen at room temperature for 16 hours then pour it into cold water and extract the mixture with chloroform. Wash the combined chloroform extracts with water then concentrate under reduced pressure to a residue of substantially methyl D-nor-4,l6(20)-pregnadiene-3- one-21-oate. Purify the residue by chromotography on Florisil eluting with hexane-ether-acetone mixtures. Combine like fractions on the basis of ultraviolet and infrared spectra, retaining those fractions wherein the infrared spectra indicates the presence of an ester group and whose ultraviolet spectra indicates the presence of two conjugated systems. Combine the selected fractions and concentrate to a residue and crystallize from acetone-hexane.

o. METHYL D-NOR5,16(20)-PREGNADIENE-3ONE-21- OATE 3-ETHYLENE KETAL To a solution of 1.5 g. of methyl D-nor-4-,l6(20)-pregnadiene-3-one-21-oate in ml. of benzene add 7.5 ml. of ethylene glycol and 0.15 g. of p-toluenesulfonic acid. Stir the mixture at reflux temperature for 6 hours then cool and wash with 100 ml. of 1% aqueous sodium bicarbonate. Put the washed benzene solution on a column of Florisil and elute with ether. Combine the fractions and evaporate to a residue containing substantially methyl D-nor-5,16(20) pregnadiene 3 one 21 oate 3-ethylene ketal. Purify by crystallization from acetonehexane.

D. D- NOR-5,16 (20) PREGNADIEliE-2l-OL-3-ONE 3-ETHYLENE KETAL Add dropwise with stirring a solution of 1 g. of methyl D-n-or,5,16(20)-pregnadiene-3-one-21 oate 3 ethylene ketal in 50 ml. of freshly distilled tetrahyclrofuran, to a suspension of 500 mg. of lithium aluminum hydride in 25 ml. of tetrahydrofuran. Heat the mixture at reflux temperature for 1 hour then chill and add dropwise 10 ml. of ethyl acetate followed by 5 ml. of a saturated aqueout solution of sodium sulfate. Finally add some solid sodium sulfate and filter the mixture. Remove the solvent from the filtrate under reduced pressure to a residue of substantially D-nor-S,16(20)-pregnadiene-21-ol 3 one 3-ethylene ketal. Purify by crystallization with ether.

E. DnNOR-4,16 (20)-PREGNADIENE21-OL-3-ONE To a solution of 500 mg. of D-nor-4,16(20)-pregnadiene-21-ol-3-one 3-ethylene ketal in 40 ml. of 80% aqueous acetone add 0.5 ml. of sulfuric acid. Keep the mixture at room temperature for 18 hours then add aqueous sodium bicarbonate until the solution is basic. Concentrate the basic solution to remove most of the acetone then add Water. The resultant precipitate is filtered, washed with water, and dried giving substantially D-nor-4,16(20)- pregnadiene-21-ol-3-one. Purify by crystallization from acetone-hexane.

11 F. D-NOR-ehlGKZO)-PREGNADIENE21-OL-3-ONE 2l-ACETATE To a solution of 2 g. of D-nor-4,16(20)-pregnadiene- 2l-ol-3-one in 10 ml. of pyridine add. 2 ml. of acetic anhydride and keep the mixture at room temperature overnight, then pour the mixture into ice water and stir for 30 minutes. Filter the resulting precipitate of substantially D-nor-4,16(20)-pregnadiene-21-ol-3-one 21-acetate then washwith water and dry. Purify by crystallization from acetone-hexane.

G. D-NOR-I-PREGNENE-l6a,21 DIOL-13,20-DIONE 21-ACETAIE Toa solution of 500 mg. of D-nor-4,16(20)-pregnadiene-21-ol-3-one 21-acetate in 50 ml. of dry t-butanol add 0.3 ml. of pyridine and 1.2 ml. of t-butanol containing 11 mg. of osmium tetroxide. To this mixture add dropwise with stirring 4.8 ml. of an 0.82 N solution of hydrogen peroxide in dry t-butanol. Keep the reaction mixture at room temperature for 5 hours then bubble nitrogen through the solution for 15 minutes followed by the addition of 600 mg. of sodium sulfite in 30 ml. of water. After 5 minutes neutralize the mixture with 10% acetic acid and dilute'with 200 ml. of water then extract with chloroform. Wash the combined extracts with water, then concentrate to a residue under reduced pressure. Dissolve the residue in a mixture of 5 ml. of pyridine and 1 ml. of

acetic anhydride and allow to stand overnight. Pour the mixture into ice water and filter the resulting precipitate of substantially D-nor-4-pregnene-16a,21-diol-3,20-dione ZI-acetate. Purify by crystallization from acetone-hexane.

H. D-NOR -1-PREGNENE-l6a,2l-DIOL-3,20-DIONE D-Nr-4-pregnene-16a-0l-3,20-di0ne and 16 esters thereof A. D-xon-4-PrmGunman-m21-DroL 3,20-D1ox1s 21p-TOLUENESULFONATE To a solution of l g. of D-nor-4-pregnene-16a,2l-diol- 3,20-dione in 20 ml. of pyridinechilled to -20 C. add a solution of 1 g. of p-toluenesulfonyl chloride in 5 ml. of methylene chloride. Stir the mixture at 20' C. overnight then add a few pieces of ice to the reaction mixture and dilutewith methylene chloride. Wash the organic solution with water, dilute hydrochloric acid, sodium bicarbonate, then water, then dry over magnesium sulfate and concentrate under reduced pressure to a residue of substantially D-nor-4-pregnene-16a,21-diol-3,20-dione 21- p-toluenesulfonate. Purify by crystallization from acetonehexane.

B. D-NOR-d-PREGNENE-l6a-OL-3,20DIONE Warm a solution of 300 mg. of D-nor-4-pregnene-l6a, 21-diol-3,20.-dione 21-p'toluenesulfonate in 10 ml. of acetone then add a warm solution of 1 g. of sodium iodide in 5 ml. of acetone. Warm the mixture on a steam bath for minutes then add 1 ml. of acetic acid and heat the mixture an additional 15 minutes. Add sodium bisulfite solution until the color of the reaction mixture is discharged, then dilute with water. Filter the resulting precipitate of substantially D-nor-4-pregnene-16a-ol-3,20- dione and wash with Water and dry. Purify by crystallize tion from acetone-hexane.

C. D-NOR--PREGNENE-lfia-OL3,20DIONE 16-ACETATE Heat a solution of 2 g. of D-nor-4-pregnene-16a-ol- 3,20-dione in a mixture of 20 ml. of acetic acid and 4 12 ml. of trifluoroacetic anhydride on the steam bath under anhydrous conditions for 1 hour. Pour the reaction mixture into ice Water and filter the resulting precipitate of substantially D-nor-4-pregnene-1 6a-ol-3,20-dione 16-ace' tate. Purify by crystallization from acetone-hexane.

D. D-NOR-4-PREGNENE-16a.-OL-3,20-DIONE 16 CAPROAI'E Heat a solution of 500 mg. of D-nor-4-pregnene-16a-ol- 3,20-dione in a mixture of 5 ml. of caproic acid and 1 ml. of trifiuoroacetic anhydride at C. for 45 minutes. Pour the mixture into ice Water and bring to neutrality with sodium carbonate. Extract the mixture with methyl ene chloride. Wash the combined extracts with water, dry over magnesium sulfate then concentrate to a residue of substantially D-nor-4-pregnene-16a-ol-3,20-dione 16-caproate. Purify by crystallization from ether.

EXAMPLE 7 D-nor-I,4-pregnadiene-16a-0l-3,20-di0ne and the 16-acetate ester thereof Subject D-nor-4-pregnene-16a-ol-3,20-dione to the ac tion of a. culture of Corynebacterium simplex in the manner described in Example 3. The resultant product is isolated in the described manner and purified by crystalliza tion from acetone-hexane yielding D-nor-1,4-pregnadiene- 16ot-0l-3,2QCii01'16.

B. D NOR 1,i-PREGNADIENE-16a-OL-3,20-DIONE 16-ACETA'1E Acetylate D-nor-l,4-pregnadiene-16a-ol-3,20-dione with acetic acid and trifiuoroacetic anhydride in the manner described in Example 6C. Isolate the resultant product in the described manner and purify by crystallization from acetonerhexane giving D-nor-1,4-pregnadiene-16a-ol-3,20- dione 16-acetate.

EXAMPLE 8 means of acetic anhydride in pyridine to form lfi-diazo.

19-nor-5-androstene-3 3-01- 17 -one 3-acetate.

In a manner similar to that described in Example 23, irradiate 16 diazo-19-nor-5-androstene-3fl-ol-17-one 3- acetate in a dioxanewater solution with ultraviolet light to give D nor-19-nor-5-androstene-313-ol-16/9-carboxylic.

acid 3-acetate.

B. D-NOR-1D-NOR'S-ANDROSTENE-35-OL1(SB-CAR- BOXYLIC ACID CHLORIDE 3-ACETATE In a manner similar to thatdescribed in Example 1A. treat D nor 19-nor-5-androstene-3fi-ol-l6fi carboxylic acid 3-acetate with oxalyl chloride in benzene and isolate the resultant product in the described manner to give D- nor-19-nor-5-androstene-3 S-ol-16/9-carboxylic acid chloride 3-acetate.

In a manner similar to that described in the alternative procedure of Example 1C, treat D-nor-19nor-5- androstene-3fi-ol-16fl-carboxylic acid chloride 3-acetate with dimethyl cadmium inbenzene. Isolate the resultant product in the described manner to give D-nor-l9-nor-5- pregnene-3 8-ol-20-one 3-acetate.

Isolate the resultant product in the described manner 5 to give D-nor-S-pregnene-3fi-ol-20-one.

E. D-NOR-lQ NOR-LPREGNENE-li,2O-DIONE (D-NOR-19- NOR-PROGESTERONE) In a manner similar to that described in Example 1E, treat D-nor-10-nor-5-pregnene-3fi-ol-20-one with chromic acid/sulfuric acid reagent. Then isolate and purify the resultant .product in the described manner to give D-nor- 19-nor-progesterone.

I claim:

1. A member selected from the group consisting of 16-X-D-nor-progesterone, wherein X is a member selected from the group consisting of hydrogen, hydroxy, and lower alkanolyloxy, and l-dehydro analogs thereof.

2. A compound according to claim 1 wherein X is hydrogen, said compound being D-nor-progesterone.

References Cited Fieser et al.: Steroids, pp. 589 and 590 (1959).

Stecher et al.: The Merck Index, p. 856 (1960) seventh edition.

Cava et al.: J. Am. Chem. Soc., vol. 84, pp. 115 and 116 (1962).

Meinwald et al.: I. Am. Chem. Soc., vol. 84, pp. 116 and 117 (1962).

Mateos et al.: Boletin Del Institute Quimica, U.N.A.M., vol. 13, pp. 3-5 (1961).

15 LEON ZITVER, Primary Examiner.

M. JACOB, Assistant Examiner.

UNITED STATES PATENT OFFICE CERTIFICATE OF CORRECTION Patent No. 3,352,920 November 14, 1967 Jerrold Meinwald It is hereby certified that error appears in the above numbered patent requiring correction and that the said Letters Patent should read as corrected below.

Column 2, lines 59 to 64, for that portion of formula V reading "-C0O" read --COOH column 5, line 37 for "loom-oxide" read l6a-oxido column 7, line 10, for "21- ddiazo" read. -l 2l-diazo column 10., lines 58 and S9, for "aqueout" read. aqueous line, 66,for "D-nor4,l6" read D-nor-S,l6 column 13, line 10, for "D-nor-lO-nor" read D-nor-l9-nor Signed and sealed this 31st day of December 1968.

SEAL) mast:

EDWARD J. BRENNER Commissioner of Patents Edward M. Fletcher, Jr.

\ttesting Officer 

1. A MEMBER SELECTED FROM THE GROUP CONSISTING OF 16-X-D-NOR-PROGESTERONE, WHEREIN X IS A MEMBER SELECTED FROM THE GROUP CONSISTING OF HYDROGEN, HYDROXY, AND LOWER ALKANOLYLOXY, AND 1-DEHYDRO ANALOGS THEREOF. 